Open Access

Epidemiology of vampire bat-transmitted rabies virus in Goiás, central Brazil: re-evaluation based on G-L intergenic region

  • Shinji Hirano1,
  • Takuya Itou1Email author,
  • Adolorata AB Carvalho2,
  • Fumio H Ito3 and
  • Takeo Sakai1
BMC Research Notes20103:288

DOI: 10.1186/1756-0500-3-288

Received: 26 May 2010

Accepted: 8 November 2010

Published: 8 November 2010

Abstract

Background

Vampire bat related rabies harms both livestock industry and public health sector in central Brazil. The geographical distributions of vampire bat-transmitted rabies virus variants are delimited by mountain chains. These findings were elucidated by analyzing a high conserved nucleoprotein gene. This study aims to elucidate the detailed epidemiological characters of vampire bat-transmitted rabies virus by phylogenetic methods based on 619-nt sequence including unconserved G-L intergenic region.

Findings

The vampire bat-transmitted rabies virus isolates divided into 8 phylogenetic lineages in the previous nucleoprotein gene analysis were divided into 10 phylogenetic lineages with significant bootstrap values. The distributions of most variants were reconfirmed to be delimited by mountain chains. Furthermore, variants in undulating areas have narrow distributions and are apparently separated by mountain ridges.

Conclusions

This study demonstrates that the 619-nt sequence including G-L intergenic region is more useful for a state-level phylogenetic analysis of rabies virus than the partial nucleoprotein gene, and simultaneously that the distribution of vampire bat-transmitted RABV variants tends to be separated not only by mountain chains but also by mountain ridges, thus suggesting that the diversity of vampire bat-transmitted RABV variants was delimited by geographical undulations.

Background

Rabies is a zoonosis that kills infected mammals, including humans, and is mainly transmitted by carnivores. In the Americas, chiropterans (insectivorous, frugivorous and hematophagous bat) are another reservoir of this disease. Although dog-transmitted rabies in central Brazil has been reduced by aggressive vaccination programs [1], chiroptera (particularly the common vampire bat, Desmodus rotundas)-transmitted rabies remains endemic in this region, and harms both the livestock industry and the public health sector [2, 3].

To date, vampire bat-transmitted rabies in livestock has been controlled by reducing the population of vampire bats and by vaccinating livestock [3, 4]. However, the depopulation of vampire bats has limitations and the effects are temporary, while vaccination of livestock is only carried out for some animals and is ineffective in decreasing rabies levels in vampire bats.

For the sustainable and effective control of vampire bat rabies, further knowledge of epidemiological features, such as vampire bat ecology and the dynamics of vampire bat-transmitted rabies, is necessary. Molecular epidemiological analysis of vampire bat-transmitted cattle rabies cases using the partial nucleoprotein gene, which is the most conserved gene in the rabies virus (RABV) genome, has suggested that the distribution of variants in Brazil is delimited by mountain chains and clustered in tens of thousands of square kilometers [5]. However, the vampire bats migrate several kilometers from their nests [6]. To elucidate a more detailed genetic clustering and geo-distribution of genetic clades of vampire bat-transmitted RABV, 204 isolates from Goiás, which includes the 185 isolates analyzed previously, were employed and analyzed by a phylogenetic method based on a nucleotide sequence encompassing the G-L intergenic region locating between glycoprotein (G) and polymerase (L) gene loci, which is the most divergent region in the RABV genome and is used for monitoring epidemiological changes in the evolution of RABV [7, 8].

Results

Phylogenetic analysis

The 204 RABV isolates divided into 8 phylogenetic lineages in the previous nucleoprotein gene analysis were divided into 10 phylogenetic lineages with significant bootstrap values (Figure 1; details shown in Table 1). Isolates of the C-5 and C-6 lineages designated by Kobayashi et al. belonged to the A-lineage, while the B-lineage consisted of some isolates of C-5. The C-, D-, F- and J-lineages included isolates belonging to C-12, C-1, C-22 and C-3, respectively. Isolates of C-20 belonged to the I-lineage, and C-21 was divided into three lineages; G-, H- and I-lineages. Two isolates not belonging to any lineages in previous studies were assigned to the E-lineage.
https://static-content.springer.com/image/art%3A10.1186%2F1756-0500-3-288/MediaObjects/13104_2010_Article_692_Fig1_HTML.jpg
Figure 1

NJ tree based on 619-nt sequence of the partial G and the G-L intergenic region. Full circles on internal branches indicate significant bootstrap values (>70%). The number of isolates retaining 100% nucleotide similarity with the representative isolate is shown in parentheses. Lineages of the isolates defined in a previous study [5] are shown in italics after a diagonal (Ud: undefined).

Table 1

Isolates from Goiás

    

Grouping

Accession No.

Sample

Species

Location

Year

This study

Previous studyc

N203-nt

Partial G & GL

BRbv371

Cattle

Caldas Novas

2002

A

C-5

AB307182

AB544082

BRbv372

Cattle

Água Limpa

2002

A

C-5

AB307183

AB544083

BRbv373

Cattle

Caldas Novas

2002

A

C-5

AB307184

AB544084

BRbv375

Cattle

Santa Cruz de Goias

2002

A

C-5

AB307186

AB544085

BRbv379

Cattle

Corumbaíba

2002

A

C-5

AB307190

AB544089

BRbv380

Cattle

Corumbaíba

2002

A

C-5

AB307191

AB544090

BRbv381

Cattle

Cristalina

2002

A

C-5

AB307192

AB544091

BRbv386

Cattle

Ipameri

2002

A

C-5

AB307197

AB544095

BRbv391

Cattle

Caldas Novas

2002

A

C-5

AB307201

AB544099

BRbv392

Cattle

Água Limpa

2002

A

C-5

AB307202

AB544100

BRbv393

Cattle

Itumbiara

2002

A

C-5

AB307203

AB544101

BRbv395

Cattle

Itumbiara

2002

A

C-5

AB307205

AB544102

BRbv396

Cattle

Itumbiara

2002

A

C-5

AB307206

AB544103

BRbv397

Cattle

Buriti Alegre

2002

A

C-5

AB307207

AB544104

BRbv398

Cattle

Itumbiara

2002

A

C-5

AB307208

AB544105

BRbv400

Cattle

Buriti Alegre

2002

A

C-5

AB307210

AB544107

BRbv404

Cattle

Buriti Alegre

2002

A

C-5

AB307214

AB544111

BRbv405

Cattle

Buriti Alegre

2002

A

C-5

AB307215

AB544112

BRbv407

Cattle

Goiandira

2002

A

C-5

AB307217

AB544114

BRbv412

Cattle

Itapaci

2002

A

C-5

AB307222

AB544118

BRbv413

Cattle

Caldas Novas

2002

A

C-5

AB307223

AB544119

BRbv416

Cattle

Morrinhos

2002

A

C-5

AB307226

AB544122

BRbv421

Cattle

Nova Aurora

2002

A

C-5

AB307231

AB544124

BRbv429

Cattle

Corumbaíba

2002

A

Uda

AB307238

AB544129

BRbv432

Cattle

Ipameri

2002

A

C-5

AB307241

AB544132

BRbv438

Cattle

Buriti Alegre

2002

A

C-5

AB307247

AB544137

BRhr441

Horse

Buriti Alegre

2002

A

Ud

AB307251

AB544140

BRbv442

Cattle

Buriti Alegre

2002

A

C-5

AB307252

AB544141

BRbv447

Cattle

Urutaí

2002

A

C-6

AB307255

AB544143

BRbv449

Cattle

Caldas Novas

2002

A

C-5

AB307256

AB544145

BRbv451

Cattle

Ipameri

2002

A

C-5

AB307258

AB544147

BRbv453

Cattle

São Luis de Montes Belos

2002

A

C-5

AB307260

AB544149

BRbv456

Cattle

Orizona

2002

A

Ud

AB307263

AB544150

BRbv457

Cattle

Água Limpa

2002

A

C-5

AB307264

AB544151

BRbv458

Cattle

Buriti Alegre

2002

A

C-5

AB307265

AB544152

BRbv466

Cattle

Itumbiara

2002

A

C-5

AB307273

AB544157

BRbv469

Cattle

Ipameri

2002

A

C-6

AB307275

AB544159

BRbv471

Cattle

Santa Cruz de Goias

2002

A

C-6

AB307277

AB544161

BRbv472

Cattle

Ipameri

2002

A

C-6

AB307278

AB544162

BRbv473

Cattle

Itumbiara

2002

A

C-5

AB307279

AB544163

BRbv475

Cattle

Caldas Novas

2002

A

C-5

AB307281

AB544165

BRbv477

Cattle

Itumbiara

2002

A

C-5

AB307283

AB544166

BRbv478

Cattle

Itumbiara

2002

A

C-5

AB307284

AB544167

BRbv482

Cattle

Panamá

2002

A

C-5

AB307288

AB544170

BRhr483

Horse

Panamá

2002

A

Ud

AB307291

AB544171

BRbv486

Cattle

Itumbiara

2002

A

C-5

AB307293

AB544172

BRbv489

Cattle

Itumbiara

2002

A

Ud

AB307295

AB544174

BRbv493

Cattle

Ipameri

2002

A

C-6

AB307298

AB544177

BRbv495

Cattle

Panamá

2002

A

C-5

AB307300

AB544179

BRbv496

Cattle

Panamá

2002

A

C-5

AB307301

AB544180

BRbv497

Cattle

Itumbiara

2002

A

C-5

AB307302

AB544181

BRbv498

Cattle

Bela Vista de Goiás

2002

A

C-5

AB307303

AB544182

BRbv502

Cattle

Itumbiara

2002

A

C-5

AB307306

AB544186

BRbv503

Cattle

Orizona

2002

A

Ud

AB307307

AB544187

BRbv509

Cattle

Buriti Alegre

2002

A

C-5

AB307311

AB544192

BRbv514

Cattle

Panamá

2002

A

C-5

AB307316

AB544196

BRbv516

Cattle

Orizona

2002

A

Ud

AB307318

AB544198

BRbv521

Cattle

Urutaí

2002

A

Ndb

Nd

AB544201

BRbv524

Cattle

Urutaí

2002

A

C-6

AB307323

AB544204

BRbv526

Cattle

Panamá

2002

A

C-5

AB307325

AB544206

BRbv527

Cattle

Campos Belos

2002

A

C-5

AB307326

AB544207

BRbv684

Cattle

Ipameri

2001

A

C-5

AB307412

AB544211

BRbv685

Cattle

Corumbaíba

2001

A

Nd

Nd

AB544212

BRbv686

Cattle

Ipameri

2001

A

C-5

AB307413

AB544213

BRbv687

Cattle

Corumbaíba

2001

A

C-5

AB307414

AB544214

BRbv690

Cattle

Ipameri

2001

A

C-5

AB307417

AB544215

BRbv691

Cattle

Ipameri

2001

A

C-5

AB307418

AB544216

BRbv694

Cattle

Iporá

2001

A

Nd

Nd

AB544219

BRbv697

Cattle

Campo Alegre de Goiás

2001

A

C-5

AB307423

AB544221

BRbv698

Cattle

Campo Alegre de Goiás

2001

A

C-5

AB307424

AB544222

BRbv699

Cattle

Campo Alegre de Goiás

2001

A

C-5

AB307425

AB544223

BRbv700

Cattle

Caldas Novas

2001

A

Nd

Nd

AB544224

BRbv701

Cattle

Caldas Novas

2001

A

C-5

AB307426

AB544225

BRhr704

Horse

Corumbaíba

2001

A

Nd

Nd

AB544227

BRbv705

Cattle

Ipameri

2001

A

C-5

AB307428

AB544228

BRbv707

Cattle

Corumbaíba

2001

A

Nd

Nd

AB544229

BRsp708

Sheep

Ouvidor

2001

A

Nd

Nd

AB544230

BRbv710

Cattle

Morrinhos

2001

A

C-5

AB307431

AB544232

BRbv711

Cattle

Morrinhos

2001

A

C-5

AB307432

AB544233

BRbv717

Cattle

Ipameri

2001

A

C-5

AB307438

AB544235

BRbv718

Cattle

Ipameri

2001

A

C-5

AB307439

AB544236

BRbv741

Cattle

Marzagão

2001

A

C-5

AB307450

AB544244

BRbv742

Cattle

Ipameri

2001

A

C-5

AB307451

AB544245

BRbv745

Cattle

Buriti Alegre

2001

A

C-5

AB307454

AB544248

BRbv747

Cattle

Corumbaíba

2001

A

Ud

AB307456

AB544250

BRbv748

Cattle

Campo Alegre de Goiás

2001

A

C-5

AB307457

AB544251

BRbv752

Cattle

Catalão

2001

A

Ud

AB307461

AB544254

BRhr755

Horse

Cumari

2001

A

Nd

Nd

AB544257

BRbv770

Cattle

Ipameri

2002

A

C-5

AB307476

AB544269

BRbv771

Cattle

Caldas Novas

2002

A

C-5

AB307477

AB544270

BRbv780

Cattle

Nova Crixás

2002

A

C-5

AB307484

AB544274

BRbv785

Cattle

Corumbaíba

2002

A

C-5

AB307486

AB544277

BRbv786

Cattle

Buriti Alegre

2002

A

Ud

AB307487

AB544278

BRbv796

Cattle

Pires do Rio

2002

A

C-6

AB307495

AB544283

BRbv797

Cattle

Cristalina

2002

A

C-6

AB307496

AB544284

BRbv402

Cattle

Divinópolis de Goiás

2002

B

C-5

AB307212

AB544109

BRbv406

Cattle

Posse

2002

B

C-5

AB307216

AB544113

BRbv459

Cattle

Monte Alegre de Goiás

2002

B

C-5

AB307266

AB544153

BRbv461

Cattle

Monte Alegre de Goiás

2002

B

C-5

AB307268

AB544155

BRbv462

Cattle

Monte Alegre de Goiás

2002

B

C-5

AB307269

AB544156

BRbv479

Cattle

Colinas do Sul

2002

B

C-5

AB307285

AB544168

BRbv481

Cattle

Serranópolis

2002

B

C-5

AB307287

AB544169

BRbv490

Cattle

Colinas do Sul

2002

B

C-5

AB307296

AB544175

BRbv494

Cattle

Divinópolis de Goiás

2002

B

C-5

AB307299

AB544178

BRbv500

Cattle

Colinas do Sul

2002

B

C-5

AB307304

AB544184

BRbv505

Cattle

Colinas do Sul

2002

B

C-5

AB307309

AB544188

BRhr506

Horse

Colinas do Sul

2002

B

Nd

Nd

AB544189

BRhr507

Horse

Colinas do Sul

2002

B

Nd

Nd

AB544190

BRbv510

Cattle

Monte Alegre de Goiás

2002

B

C-5

AB307312

AB544193

BRbv511

Cattle

Divinópolis de Goiás

2002

B

C-5

AB307313

AB544194

BRbv518

Cattle

Campos Belos

2002

B

C-5

AB307319

AB544199

BRbv525

Cattle

Panamá

2002

B

C-5

AB307324

AB544205

BRbv753

Cattle

Monte Alegre de Goiás

2001

B

C-5

AB307462

AB544255

BRbv762

Cattle

Divinópolis de Goiás

2001

B

C-5

AB307470

AB544263

BRbv387

Cattle

Doverlândia

2002

C

C-12

AB307198

AB544096

BRbv401

Cattle

Mineiros

2002

C

C-12

AB307211

AB544108

BRbv424

Cattle

Doverlândia

2002

C

C-12

AB307234

AB544125

BRbv692

Cattle

Mineiros

2001

C

C-12

AB307419

AB544217

BRbv712

Cattle

Doverlândia

2001

C

C-12

AB307433

AB544234

BRbv751

Cattle

Mineiros

2001

C

C-12

AB307460

AB544253

BRbv383

Cattle

Nova América

2002

D

C-1

AB307194

AB544092

BRbv384

Cattle

Nova América

2002

D

C-1

AB307195

AB544093

BRbv409

Cattle

Rubiataba

2002

D

C-1

AB307219

AB544115

BRbv411

Cattle

Morrinhos

2002

D

C-1

AB307221

AB544117

BRbv452

Cattle

Rubiataba

2002

D

C-1

AB307259

AB544148

BRbv767

Cattle

Rubiataba

2002

D

C-1

AB307474

AB544267

BRbv793

Cattle

Urutaí

2002

D

C-1

AB307493

AB544282

BRbv440

Cattle

Nova Crixás

2002

E

Ud

AB307249

AB544139

BRbv519

Cattle

Mundo Novo

2002

E

Ud

AB307320

AB544200

BRhr389

Horse

Nova Crixás

2002

F

Nd

Nd

AB544098

BRbv434

Cattle

Itapuranga

2002

F

Ud

AB307243

AB544133

BRbv439

Cattle

Mundo Novo

2002

F

C-22

AB307248

AB544138

BRbv488

Cattle

Carmo do Rio Verde

2002

F

Ud

AB307294

AB544173

BRbv749

Cattle

Nova Crixás

2001

F

C-22

AB307458

AB544252

BRbv757

Cattle

Monte Alegre de Goiás

2001

F

C-22

AB307465

AB544259

BRbv763

Cattle

Aruanã

2002

F

C-22

AB307471

AB544264

BRbv773

Cattle

Aruanã

2002

F

C-22

AB307479

AB544272

BRbv774

Cattle

Mozarlândia

2002

F

C-22

AB307480

AB544273

BRsp787

Sheep

Ipameri

2002

F

Nd

Nd

AB544279

BRbv378

Cattle

Novo Brasil

2002

G

C-21

AB307189

AB544088

BRbv425

Cattle

Novo Brasil

2002

G

C-21

AB307235

AB544126

BRbv445

Cattle

Morrinhos

2002

G

Ud

AB307254

AB544142

BRbv448

Cattle

Buriti de Goiás

2002

G

C-21

AB307256

AB544144

BRhr467

Horse

Buriti de Goiás

2002

G

Nd

Nd

AB544158

BRbv474

Cattle

Moiporá

2002

G

C-21

AB307280

AB544164

BRbv501

Cattle

Anicuns

2002

G

C-21

AB307305

AB544185

BRbv515

Cattle

Córrego do Ouro

2002

G

C-21

AB307317

AB544197

BRbv522

Cattle

Buriti de Goiás

2002

G

Nd

Nd

AB544202

BRbv523

Cattle

Córrego do Ouro

2002

G

C-21

AB307322

AB544203

BRbv764

Cattle

Córrego do Ouro

2002

G

C-21

AB307472

AB544265

BRbv385

Cattle

Ivolândia

2002

H

C-21

AB307196

AB544094

BRbv414

Cattle

Ivolândia

2002

H

C-21

AB307224

AB544120

BRbv415

Cattle

Ivolândia

2002

H

C-21

AB307225

AB544121

BRbv426

Cattle

Amorinópolis

2002

H

C-21

AB307236

AB544127

BRbv427

Cattle

Ivolândia

2002

H

C-21

AB307237

AB544128

BRbv431

Cattle

Palestina de Goiás

2002

H

Ud

AB307240

AB544131

BRbv437

Cattle

Goiandira

2002

H

Ud

AB307246

AB544136

BRbv693

Cattle

Amorinópolis

2001

H

C-21

AB307420

AB544218

BRhr703

Horse

Ivolândia

2001

H

Nd

Nd

AB544226

BRbv736

Cattle

Amorinópolis

2001

H

C-21

AB307446

AB544242

BRbv744

Cattle

Amorinópolis

2001

H

C-21

AB307453

AB544247

BRbv370

Cattle

Caiapônia

2002

I

C-21

AB307181

AB544081

BRbv376

Cattle

Caiapônia

2002

I

C-21

AB307187

AB544086

BRbv377

Cattle

Bom Jardim de Goiás

2002

I

C-21

AB307188

AB544087

BRbv388

Cattle

Piranhas

2002

I

Ud

AB307199

AB544097

BRbv399

Cattle

Rio Verde

2002

I

C-21

AB307209

AB544106

BRbv403

Cattle

Piranhas

2002

I

C-20

AB307213

AB544110

BRbv410

Cattle

Piranhas

2002

I

C-21

AB307220

AB544116

BRbv420

Cattle

Piranhas

2002

I

C-21

AB307230

AB544123

BRbv430

Cattle

Buriti Alegre

2002

I

Ud

AB307239

AB544130

BRbv435

Cattle

Bom Jardim de Goiás

2002

I

C-20

AB307244

AB544134

BRbv436

Cattle

Piranhas

2002

I

Ud

AB307245

AB544135

BRbv450

Cattle

Palestina de Goiás

2002

I

C-21

AB307257

AB544146

BRbv460

Cattle

Caiapônia

2002

I

C-20

AB307267

AB544154

BRbv470

Cattle

Piranhas

2002

I

C-20

AB307276

AB544160

BRbv491

Cattle

Caiapônia

2002

I

C-20

AB307297

AB544176

BRbv499

Cattle

Bom Jardim de Goiás

2002

I

Nd

Nd

AB544183

BRbv513

Cattle

Caiapônia

2002

I

C-20

AB307315

AB544195

BRbv681

Cattle

Piranhas

2001

I

Nd

Nd

AB544208

BRbv682

Cattle

Piranhas

2001

I

Nd

Nd

AB544209

BRbv683

Cattle

Piranhas

2001

I

C-21

AB307411

AB544210

BRbv696

Cattle

Piranhas

2001

I

C-21

AB307422

AB544220

BRbv709

Cattle

Piranhas

2001

I

C-21

AB307430

AB544231

BRbv720

Cattle

Arenópolis

2001

I

C-21

AB307440

AB544237

BRbv721

Cattle

Arenópolis

2001

I

C-21

AB307441

AB544238

BRbv722

Cattle

Arenópolis

2001

I

C-21

AB307442

AB544239

BRbv734

Cattle

Arenópolis

2001

I

C-21

AB307444

AB544240

BRbv735

Cattle

Arenópolis

2001

I

Ud

AB307445

AB544241

BRbv738

Cattle

Piranhas

2001

I

C-20

AB307448

AB544243

BRbv743

Cattle

Marzagão

2001

I

C-20

AB307452

AB544246

BRbv746

Cattle

Caiapônia

2001

I

C-20

AB307455

AB544249

BRbv756

Cattle

Arenópolis

2001

I

C-21

AB307464

AB544258

BRbv758

Cattle

Iporá

2001

I

C-21

AB307466

AB544260

BRbv759

Cattle

Diorama

2001

I

C-21

AB307467

AB544261

BRbv760

Cattle

Palestina de Goiás

2001

I

C-21

AB307468

AB544262

BRbv765

Cattle

Caiapônia

2002

I

C-20

AB307473

AB544266

BRbv769

Cattle

Palestina de Goiás

2002

I

C-21

AB307475

AB544268

BRbv772

Cattle

Arenópolis

2002

I

C-21

AB307478

AB544271

BRhr782

Horse

Palestina de Goiás

2002

I

Nd

Nd

AB544275

BRbv784

Cattle

Piranhas

2002

I

C-20

AB307485

AB544276

BRbv790

Cattle

Pires do Rio

2002

I

C-21

AB307490

AB544280

BRbv508

Cattle

Porangatatu

2002

J

C-3

AB307310

AB544191

BRbv754

Cattle

Alvorada do Norte

2001

J

C-3

AB307463

AB544256

BRbv792

Cattle

Morrinhos

2002

J

C-3

AB307492

AB544281

aUndefined; bNo data; cKobayashi et al. (2008)

Geographical plotting

In this study, geographical areas in Goiás were divided by mountain chains into the Northwest, North central, Northeast and South regions (Figure 2). Most isolates of the A-lineage were distributed in the South region. The B-lineage was likely to exist in the Northeast and North central regions. The isolates belonging to the C-, E-, F-, G-, H- and I-lineages were distributed in the Northwest region. The G-, H- and I-lineages have narrow distributions and are apparently separated by mountain ridges (Figure 2; Area I), while the A-lineage was distributed in a wide range throughout a Southeastern basin (Figure 2; Area II). The isolates of the D-lineage were plotted on an eastern edge of the North central region. The J-lineage had a wide geographical distribution in Goiás.
https://static-content.springer.com/image/art%3A10.1186%2F1756-0500-3-288/MediaObjects/13104_2010_Article_692_Fig2_HTML.jpg
Figure 2

Distribution of vampire bat-transmitted RABV isolates in Goiás. Area I: distribution of G-, H- and I-lineages in mountain ridges. Area II: distribution of A-lineage in the basin. Symbols are same as those used in Fig. 1. Numbers inside the symbols indicate the number of isolates obtained in the same region. Small and open symbols indicated that only one isolate was obtained.

Discussion

Previous studies had elucidated that the distributions of vampire bat and transmitted RABV are delimited by mountain chains [5]. In the present study, it was reconfirmed that mountain chains divide the distribution patterns of each viral lineage. Furthermore, the isolates belonging to C-5 having a wide range in Goiás were divided into the A- and B-lineages, and were found to be distributed in the South and North regions on either side of a mountain chain. This finding supports Kobayashi's hypothesis that distribution of vampire bat-transmitted RABV is affected by mountain chains.

The same variants of vampire bat-transmitted RABV were spread widely in flat low lands (< 800 m), but at higher elevations (800-1600 m), they had a narrower distribution [9]. However, the G-, H- and I-lineages were found to be separated by mountain ridges in low areas (400-800 m) located in a southern undulating area of the Northwestern region (Figure 2; Area I). Furthermore, the A-lineage was located in an eastern basin of the South region (> 800 m; Figure 2; Area II). Considering that the higher lands showed an undulating landscape, the results suggest that the distribution patterns of vampire bat-transmitted RABV variants depend on such undulations. On the other hand, the distribution of common vampire bats in a valley is limited by the ridges that form the valley [10], thus supporting the notion that the distribution of RABV variants is affected by smaller topography than mountain chains.

Conclusions

The present study analyzed the epidemiology of vampire bat-transmitted RABV using a 619-nt region containing the partial glycoprotein gene and the G-L intergenic region, and indicated that the isolates can be further divided into several phylogenetic lineages with significant bootstrap values when compared to characterization based on the 203-nt partial N gene. Furthermore, the phylogenetic lineages were divided by both mountain chains and mountain ridges. In future studies, it will be important to analyze samples from different time points and to elucidate the dynamics of vampire bat-transmitted rabies in order to establish effective and sustainable control measures for preventing rabies circulation among vampire bats.

Methods

Samples

A total of 204 samples obtained from 192 cattle, 10 horses and 2 sheep in Goiás from October 2001 to August 2002 were employed, which had been confirmed as rabies positive through fluorescent antibody test and mouse inoculation test (Table 1). Viral RNA was extracted from the brain as described previously [11]. Lineages of the 164 cattle isolates, C-1, C-3, C-5, C-6, C-12, C-20, C-21 and C-22, were previously characterized based on a 203-nt sequence of the nucleoprotein gene [5], and are shown in Table 1.

Determination of nucleotide sequences

RT-PCR and direct sequencing with the HmG5-1302 (4619TGTTGAAGTTCACCTYCCMGATGT4642, positions relative to PV strain genome (Accession No. M13215) and RVLa-1 (5435ATRGGGTCATCATAAACCTC5416) primer pair were performed as described previously [11]. The target sequence includes the partial glycoprotein gene and G-L intergenic region. Nucleotide sequences were determined using the ATGC program version 4.02 (GENETYX Co., Tokyo, Japan).

Phylogenetioc analysis

Multiple nucleotide sequence alignments of the partial glycoprotein gene and G-L intergenic region were generated by the ClustalW package in MEGA ver. 4.0 [12]. A phylogenetic tree was constructed by the neighbor joining (NJ) method with bootstrap analysis (1000 psuedoreplicates) under the p-distance model. Phylogenetic clustering supported by a bootstrap value exceeding 70% was regarded as a reliable lineage [13]. Results were validated by the maximum likelihood method using PhyML [14]. In order to reconfirm the shape of the NJ tree, the ML tree was constructed under HKY substitution model justified by MODELTEST packaged in Hyphy program [15].

Geographical plotting

The 204 RABV isolates were plotted onto a geographical map described using the DIVA-GIS program [16] with GIS data from Instituto Brasileiro de Geografia e Estatística [17] and DIVA-GIS gData [18].

Declarations

Acknowledgements

This work was partly supported by the "Strategic Research Base Development" Program for Private Universities subsidized by the Ministry of Education, Culture, Sports, Science and Technology (MEXT) of Japan.

Authors’ Affiliations

(1)
Nihon University Veterinary Research Center
(2)
Faculty of Agriculture and Veterinary Science, Department of Preventive Veterinary Medicine, UNESP, Via de Acesso Prof. Paulo Donato Castellane
(3)
Department of Preventive Veterinary Medicine and Animal Health, Faculty of Veterinary Medicine and Zootechny, University of São Paulo, Av. Prof. Dr. Orlando Marques de Paiva

References

  1. Romijn PC, van der Heide R, Cattaneo CA, Silva RC, van der Poel WH: Study of lyssaviruses of bat origin as a source of rabies for other animal species in the State of Rio De Janeiro, Brazil. Am J Trop Med Hyg. 2003, 69: 81-86.PubMedGoogle Scholar
  2. Goncalves MA, Sa Neto RJ, Brazil TK: Outbreak of aggressions and transmission of rabies in human beings by vampire bats in northeastern Brazil. Rev Soc Bras Med Trop. 2002, 35: 461-464. 10.1590/S0037-86822002000500006.PubMedView ArticleGoogle Scholar
  3. Mayen F: Haematophagous bats in Brazil, their role in rabies transmission, impact on public health, livestock industry and alternatives to an indiscriminate reduction of bat population. J Vet Med B Infect Dis Vet Public Health. 2003, 50: 469-472.PubMedView ArticleGoogle Scholar
  4. Massad E, Coutinho FA, Burattini MN, Sallum PC, Lopez LF: A mixed ectoparasite--microparasite model for bat-transmitted rabies. Theor Popul Biol. 2001, 60: 265-279. 10.1006/tpbi.2000.1494.PubMedView ArticleGoogle Scholar
  5. Kobayashi Y, Sato G, Mochizuki N, Hirano S, Itou T, Carvalho AA, Albas A, Santos HP, Ito FH, Sakai T: Molecular and geographic analyses of vampire bat-transmitted cattle rabies in central Brazil. BMC Vet Res. 2008, 4: 44-10.1186/1746-6148-4-44.PubMed CentralPubMedView ArticleGoogle Scholar
  6. Arellano-Sota C: Biology, ecology, and control of the vampire bat. Rev Infect Dis. 1988, 10: S615-619.PubMedView ArticleGoogle Scholar
  7. Nel LH, Sabeta CT, von Teichman B, Jaftha JB, Rupprecht CE, Bingham J: Mongoose rabies in southern Africa: a re-evaluation based on molecular epidemiology. Virus Res. 2005, 109: 165-173. 10.1016/j.virusres.2004.12.003.PubMedView ArticleGoogle Scholar
  8. Ngoepe CE, Sabeta C, Nel L: The spread of canine rabies into Free State province of South Africa: A molecular epidemiological characterization. Virus Res. 2009, 142: 175-180. 10.1016/j.virusres.2009.02.012.PubMedView ArticleGoogle Scholar
  9. Kobayashi Y, Ogawa A, Sato G, Sato T, Itou T, Samara SI, Carvalho AA, Nociti DP, Ito FH, Sakai T: Geographical distribution of vampire bat-related cattle rabies in Brazil. J Vet Med Sci. 2006, 68: 1097-1100. 10.1292/jvms.68.1097.PubMedView ArticleGoogle Scholar
  10. Trajano E: Movements of Cave Bats in Southeastern Brazil, with Emphasis on the Population Ecology of the Common Vampire Bat, Desmodus rotundus (Chiroptera). Biotropica. 1996, 28: 121-129. 10.2307/2388777.View ArticleGoogle Scholar
  11. Sato G, Itou T, Shoji Y, Miura Y, Mikami T, Ito M, Kurane I, Samara SI, Carvalho AA, Nociti DP, Ito FH, Sakai T: Genetic and phylogenetic analysis of glycoprotein of rabies virus isolated from several species in Brazil. J Vet Med Sci. 2004, 66: 747-753. 10.1292/jvms.66.747.PubMedView ArticleGoogle Scholar
  12. Tamura K, Dudley J, Nei M, Kumar S: MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Mol Biol Evol. 2007, 24: 1596-1599. 10.1093/molbev/msm092.PubMedView ArticleGoogle Scholar
  13. Hills DM, Bull JJ: An empirical test of boostrapping as a method for assessing confidence in phylogenetic analysis. Syst Biol. 1993, 42: 182-192.View ArticleGoogle Scholar
  14. Guindon S, Lethiec F, Duroux P, Gascuel O: PHYML Online--a web server for fast maximum likelihood-based phylogenetic inference. Nucleic Acids Res. 2005, 33: W557-559. 10.1093/nar/gki352.PubMed CentralPubMedView ArticleGoogle Scholar
  15. Pond SL, Frost SD, Muse SV: HyPhy: hypothesis testing using phylogenies. Bioinformatics. 2005, 21: 676-679. 10.1093/bioinformatics/bti079.PubMedView ArticleGoogle Scholar
  16. Hijmans RJ, Guarino L, Cruz M, Rojas E: Computer tools for spatial analysis of plant genetic resources data: 1. DIVA-GIS. Plant Genet Res Newsl. 2002, 127: 15-19.Google Scholar
  17. Instituto Brasileiro de Geografia e Estatística (IBGE). [http://www.ibge.gov.br/servidor_arquivos_geo]
  18. DIVA-GIS gData. [http://www.diva-gis.org/gData]

Copyright

© Itou et al; licensee BioMed Central Ltd. 2010

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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